RESUMO
Plants subjected to stress need to respond rapidly and efficiently to acclimatize and survive. In this paper, we investigated a selected gene set potentially involved in early cell reprogramming in two rice genotypes with contrasting salinity tolerance (Pokkali tolerant and IR29 susceptible) in order to advance knowledge of early molecular mechanisms of rice in dealing with salt stress. Selected genes were evaluated in available transcriptomic data over a short period of 24 h and involved enzymes that avoid ROS formation (AOX, UCP and PTOX), impact ATP production (PFK, ADH and COX) or relate to the antioxidant system. Higher transcript accumulation of AOX (ROS balancing), PFK and ADH (alcohol fermentation) was detected in the tolerant genotype, while the sensitive genotype revealed higher UCP and PTOX transcript levels, indicating a predominant role for early transcription of AOX and fermentation in conferring salt stress tolerance to rice. Antioxidant gene analyses supported higher oxidative stress in IR29, with transcript increases of cytosolic CAT and SOD from all cell compartments (cytoplasm, peroxisome, chloroplast and mitochondria). In contrast, Pokkali increased mRNA levels from the AsA-GSH cycle as cytosolic/mitochondrial DHAR was involved in ascorbate recovery. In addition, these responses occurred from 2 h in IR29 and 10 h in Pokkali, indicating early but ineffective antioxidant activity in the susceptible genotype. Overall, our data suggest that AOX and ADH can play a critical role during early cell reprogramming for improving salt stress tolerance by efficiently controlling ROS formation in mitochondria. We discuss our results in relation to gene engineering and editing approaches to develop salinity-tolerant crops.
RESUMO
Understanding into acerola (Malpighia emarginata) molecular and biochemical bases is still obscure, despite it is one of the most important natural source of vitamin C for humans. Recently, our research group published the first data on acerola transcriptome generating valuable information to identify reference genes for RT-qPCR in this species. Hence, this study aimed to identify the most stably expressed genes based on acerola transcriptome data, and further to evaluate the suitability of F-box, U3, Merad50-ATPase, TGD4, NOB1, PA-RNA, RCC1, RBL and PGAL candidates for accurate gene expression normalization in leaf, flower and fruit at 12, 16 and 20 days after anthesis using RT-qPCR analysis. Three algorithms, geNorm, NormFinder, and BestKeeper confirmed the expression stability of all nine candidate reference genes, whereas RefFinder consensually summarized a comprehensive gene ranking. Based on geNorm, the combination of the most stable reference genes RBL and U3 for leaf/flower group, TGD4, F-box and PGAL (fruit developmental stages or fruit/leaf), RCC1, PGAL and RBL (fruit/flower) and RCC1, RBL, TGD4 and PGAL (total samples) were required for accurate normalization. Moreover, the use of these reference genes to assess the expression profile of GMP1 and NAT3 genes confirmed the reliability of ranking and defined the best combination of genes recommended by geNorm and RefFinder. This work will benefit further RT-qPCR studies in these acerola organs by offering a foundation for accurate normalization of gene expression profiling.
Assuntos
Perfilação da Expressão Gênica/normas , Malpighiaceae/genética , Transcriptoma/genética , Algoritmos , Flores/genética , Frutas/genética , Expressão Gênica/genética , Regulação da Expressão Gênica de Plantas/genética , Genes de Plantas/genética , Folhas de Planta/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reação em Cadeia da Polimerase em Tempo Real/normas , Padrões de Referência , Reprodutibilidade dos TestesRESUMO
KEY MESSAGE: The first transcriptome coupled to metabolite analyses reveals major trends during acerola fruit ripening and shed lights on ascorbate, ethylene signalling, cellular respiration, sugar accumulation, and softening key regulatory genes. Acerola is a fast growing and ripening fruit that exhibits high amounts of ascorbate. During ripening, the fruit experience high respiratory rates leading to ascorbate depletion and a quickly fragile and perishable state. Despite its growing economic importance, understanding of its developmental metabolism remains obscure due to the absence of genomic and transcriptomic data. We performed an acerola transcriptome sequencing that generated over 600 million reads, 40,830 contigs, and provided the annotation of 25,298 unique transcripts. Overall, this study revealed the main metabolic changes that occur in the acerola ripening. This transcriptional profile linked to metabolite measurements, allowed us to focus on ascorbate, ethylene, respiration, sugar, and firmness, the major metabolism indicators for acerola quality. Our results suggest a cooperative role of several genes involved in AsA biosynthesis (PMM, GMP1 and 3, GME1 and 2, GGP1 and 2), translocation (NAT3, 4, 6 and 6-like) and recycling (MDHAR2 and DHAR1) pathways for AsA accumulation in unripe fruits. Moreover, the association of metabolites with transcript profiles provided a comprehensive understanding of ethylene signalling, respiration, sugar accumulation and softening of acerola, shedding light on promising key regulatory genes. Overall, this study provides a foundation for further examination of the functional significance of these genes to improve fruit quality traits.